A Salamander's Tail

November 18, 2009

As a graduate student, Associate Professor of Biology Janet L. Vaglia observed that a salamander's tail continues to grow throughout its life — but not in the way you might think. As humans grow, our skeletons simply become larger. Salamander bodies grow in a similar way, but their tails grow longer by adding entirely new vertebrae. The discovery led Vaglia to consider whether the continuous tail growth was connected to a salamander's ability to regenerate lost limbs and other body parts, including the tail.

"You could look at a tail that's growing throughout a salamander's life as if it's constantly regenerating, but it's not technically regeneration until you remove part of it and the healing process begins," Vaglia explains. "Growing occurs at the tip of the tail and is a linear process — it's just copying and adding on to what's already there. Regeneration that occurs when an organ is lost has a lot more work to do."

The new bones and tissue not only have to grow, but must also become functional as needed for a particular age or stage of life (you wouldn't want an adult to regenerate an immature arm). Most scientists involved in this area of research are looking for ways to heal damage to specific human parts, such as spinal cord nerves that when damaged can result in paralysis. It's easy to understand why a salamander's constantly growing tail — an extension of its spinal column — might be a good place to start to look for solutions.

In order to better understand how the regenerative and growing processes in salamanders are related, two questions have to be answered. First, Vaglia (pictured right) needs to know what types of cells are being used to build new tail segments. To find out, she uses a method called histological sectioning to examine thin slices of tissue. Because a salamander's tail grows linearly, it's possible to see cellular development over time in a single sample.

Vaglia's second research question deals with identifying the molecular signals that trigger regeneration. All vertebrates possess what are called Hox genes. These genes determine how many of each type of vertebrae should form and tell limbs where to grow. All of these instructions prevent animals from becoming a confusing mix of vertebrae and appendages in random places. So far, Hox genes are only thought to work during the embryonic stage of development. Vaglia suggests that in animals that have the ability to regenerate, Hox genes might continue to operate throughout their lives. In order to see if this is true, Vaglia's lab is cloning salamander Hox genes in order to look for their expression throughout different stages of development. This will allow her to begin deciphering some of what she calls the "chemical cocktail" responsible for continual salamander tail growth as well as regeneration.

The primary salamanders that Vaglia requires for her research can't be mail ordered from online catalogs. Each spring, she and her students travel to Hoosier National Forest to collect Southern Two-Lined salamanders. Salamanders, like many amphibians, are nocturnal, so the group will often camp overnight to collect adults that come out from hiding. Their eggs, while unable to evade capture, are just as elusive.

"The babies are hard to find because they blend in with the bottom of the stream," Vaglia says. "You have to be very still and watch for a long time. Some of my students have had quite a knack for finding them."

Back in the lab, the eggs are placed in a temperature-regulated incubator, where they hatch several weeks later. Collecting the salamanders at this early stage gives Vaglia and her students the ability to study the tail growth and regenerative processes at different points of development.

"For our regeneration studies, we can decide to amputate their tails when they hatch, or let them grow larger," Vaglia says. "We don't know if the process of growing back a tail is the same in larvae as it is in a juvenile, so it gives us a lot of different stages to look at."

When the time comes for a salamander to donate its tail to science, the process is quick and painless. Students anesthetize the salamander, then take measurements of its body and snip the tail. When the salamander awakes, it's returned to its brothers and sisters in the incubator before finally being released back into the wild. A small number of the salamanders are later put to sleep so that the base of the tail where regeneration occurs can also be studied. After a few weeks, an amputated tail will have completely regenerated.

"It's a perfect replica of the original," Vaglia says.

In spite of how strange it sounds to cut off their tails, it's a surprisingly normal part of a salamander's life. Many species of salamander have the ability to autotomize — to remove their own tail as a defensive response. For instance, if a salamander is being chased by a hungry bird, it can release connective tissue and muscular connections at certain points of its tail to cause it to fall off, leaving it behind as a wriggling countermeasure.

This year, Vaglia's salamander lab received a $200,000 grant from the National Institutes of Health, who expressed interest in research done at the undergraduate level that has implications in the medical community. While Vaglia hesitates to compare the research done in her lab with medical research, she's optimistic that the findings will make an important contribution. Vaglia said she was pleased to see the NIH reviewers acknowledge that such research is especially relevant in the ongoing search for mechanisms and factors that might help induce regrowth of tissues, such as nervous tissue.

"By uncovering the cellular and molecular mechanisms by which a salamander tail can both add vertebrae normally, as well as regenerate vertebrae, we might begin to decipher how regeneration can be stimulated in organisms that have lost the natural ability to regenerate," Vaglia says.

"What we're doing isn't going to directly solve medical problems," she adds. "This is just a piece of a puzzle that could be helpful for doing research on human injuries. If we can understand the conditions that promote tissue and organ regeneration, such work will have a major impact on human health in the future."